Universität zu Köln

Zimmermann, Jessica (2023). Interactions driving Chaperone-assisted Assembly of Proteasome Core particles. PhD thesis, Universität zu Köln.

PDF Dissertation_Jessica_Zimmermann_2023.pdf - Published Version Download (3MB)

Abstract

The ubiquitin-proteasome system (UPS) of eukaryotic cells provides an essential proteolytic control system, malfunctions of which are linked to various human diseases. The central component of the UPS is the 26S proteasome, a protease complex composed of a catalytic 20S core particle (CP) and one or two attached 19S regulatory particles (RP). Proteasome assembly is a conserved ordered process involving different intermediates and chaperones associated, which was thought to be initiated by the formation of rings comprising seven distinct α subunits. A critical intermediate in CP assembly is the 15S precursor complex (PC) containing all α and β subunits except for β7, as well as the chaperones Ump1 and Pba1-Pba2. Although the crystal structure of the mature 20S CP is already known, the step-wise process of proteasome formation is still not completely understood. The present study reveals important aspects concerning early assembly intermediates for the formation of the 15S precursor complex, the dimerization of such complexes into 20S CPs as well as specific interactions important during proteasome assembly and maturation. Using an integrated approach comprising codon adaptation and gene fusion technologies, it was possible to obtain distinct yeast proteasome components in sufficient quantities to be used for specific antibody production. In contrast to the α-ring as an early assembly intermediate, we identified two complementary complexes containing distinct subsets of α and β subunits, Complex I (α1-α4, β2-β4, Ump1) and Complex II (α5-α7, Pba3-Pba4), which constitute preliminary assembly intermediates for the formation of the 15S precursor complex, a process likely facilitated by Pba1-Pba2. With the help of in vitro binding experiments, these studies identified a novel function of the Ump1 N-terminus in the recruitment of Pro-β7 to the 15S PC complexes to drive their dimerization. Furthermore, the incorporation of the β7 subunit into the complex was confirmed to be the critical step in dimerization of 15S PCs followed by active site maturation of the catalytic subunits leading to proteolytically active 20S PCs and the degradation of Ump1.

Item Type:	Thesis (PhD thesis)
Creators:	Creators Email ORCID ORCID Put Code Zimmermann, Jessica Jessica.z@gmx.de UNSPECIFIED UNSPECIFIED
URN:	urn:nbn:de:hbz:38-722252
Date:	2023
Language:	English
Faculty:	Faculty of Mathematics and Natural Sciences
Divisions:	Faculty of Mathematics and Natural Sciences > Department of Biology > Institute for Genetics
Subjects:	Life sciences
Uncontrolled Keywords:	Keywords Language Proteasome English Assembly English Chaperone English
Date of oral exam:	20 October 2023
Referee:	Name Academic Title Dohmen, R. Jürgen Prof. Dr. Hofmann, Kay Prof. Dr.
Refereed:	Yes
URI:	http://kups.ub.uni-koeln.de/id/eprint/72225